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REVIEW ARTICLE
Middle East Respiratory Syndrome and Severe Acute RespiratorySyndrome: Current Therapeutic Options and Potential Targetsfor Novel Therapies
Julie Dyall1 • Robin Gross1 • Jason Kindrachuk2 • Reed F. Johnson3 •
Gene G. Olinger Jr.4 • Lisa E. Hensley1 • Matthew B. Frieman5 • Peter B. Jahrling1,3
Published online: 15 November 2017
� Springer International Publishing AG 2017
Abstract No specific antivirals are currently available for
two emerging infectious diseases, Middle East respiratory
syndrome (MERS) and severe acute respiratory syndrome
(SARS). A literature search was performed covering
pathogenesis, clinical features and therapeutics, clinically
developed drugs for repurposing and novel drug targets.
This review presents current knowledge on the epidemi-
ology, pathogenesis and clinical features of the SARS and
MERS coronaviruses. The rationale for and outcomes with
treatments used for SARS and MERS is discussed. The
main focus of the review is on drug development and the
potential that drugs approved for other indications provide
for repurposing. The drugs we discuss belong to a wide
range of different drug classes, such as cancer therapeutics,
antipsychotics, and antimalarials. In addition to their
activity against MERS and SARS coronaviruses, many of
these approved drugs have broad-spectrum potential and
have already been in clinical use for treating other viral
infections. A wealth of knowledge is available for these
drugs. However, the information in this review is not meant
to guide clinical decisions, and any therapeutic described
here should only be used in context of a clinical trial.
Potential targets for novel antivirals and antibodies are
discussed as well as lessons learned from treatment
development for other RNA viruses. The article concludes
with a discussion of the gaps in our knowledge and areas
for future research on emerging coronaviruses.
Key Points
The outbreaks of Middle East respiratory syndrome
(MERS) and severe acute respiratory syndrome
(SARS) were caused by emerging coronaviruses.
A variety of approaches for developing therapeutics
are discussed with emphasis on drugs that have been
approved for other indications and could be
repurposed for treating emerging coronaviral
infections.
The recent MERS and SARS outbreaks highlight the
importance of a panel of well-characterized broad-
spectrum antivirals for treating emerging viral
infections
1 Introduction
An electronic literature search for countermeasures against
Middle East respiratory syndrome coronavirus (MERS-
CoV) and severe acute respiratory syndrome coronavirus
(SARS-CoV) was performed using PubMed and Google
& Julie Dyall
1 Integrated Research Facility, Division of Clinical Research,
National Institute of Allergy and Infectious Diseases,
National Institutes of Health, Frederick, MD, USA
2 Department of Medical Microbiology, University of
Manitoba, Winnipeg, MN, Canada
3 Emerging Viral Pathogens Section, National Institute of
Allergy and Infectious Diseases, National Institutes of
Health, Frederick, MD, USA
4 University of Boston, Boston, MA, USA
5 Department of Microbiology and Immunology, University of
Maryland, School of Medicine, Baltimore, MD, USA
Drugs (2017) 77:1935–1966
https://doi.org/10.1007/s40265-017-0830-1
Scholar from 2000 through April 17, 2017. The search (key
words: Middle East Respiratory Syndrome, Severe Acute
Respiratory Syndrome, inhibitors, antivirals, therapeutics,
FDA-approved) produced 1677 citations. References
selected discussed (1) pathogenesis and history of disease,
(2) clinical countermeasures used during the 2003 SARS
and 2012 MERS outbreaks and outcomes, and (3) the
efficacy of countermeasures targeting viral components
and cellular targets of MERS-CoV and SARS-CoV. The
main emphasis was on references for drug repurposing as
an alternative to the costly development of novel drugs for
emerging coronaviral infections.
1.1 Epidemiology of MERS and SARS
Since 2003, two human coronaviruses, SARS-CoV and
MERS-CoV, emerged as global public health threats.
SARS-CoV was first identified in February 2003 in
Guangdong Province, Peoples Republic of China and was
transmitted to humans from infected civets, likely infected
from bats [1, 2]. SARS-CoV spread to 29 additional
countries and was associated with high morbidity in
humans (e.g. atypical pneumonia). Ultimately, SARS was
contained in 2004 following a highly effective public
health response but resulted in 8098 confirmed cases and
774 deaths (Fig. 1a) [3]. In 2012, MERS emerged in The
Kingdom of Saudi Arabia and presented as a severe res-
piratory disease, with frequent gastrointestinal and renal
complications. MERS-CoV, the causative agent of MERS,
was later identified as a coronavirus. MERS-CoV has
subsequently spread to 27 additional countries (Fig. 1B)
[4]. As of September 12, 2017, 2080 confirmed cases of
MERS and 722 deaths were reported [5].
Coronaviruses are enveloped, single-stranded, positive-
sense RNA viruses (Fig. 2). They are members of the
Coronavirinae subfamily of viruses and together with the
Torovirinae subfamily comprise the Coronaviridae virus
family (order Nidovirales). Coronavirinae is divided into
four genera: alpha coronavirus, beta coronavirus, gamma
coronavirus, and delta coronavirus. The coronaviruses
share a similar genome organization. The open reading
frame 1a and 1b comprise nearly 2/3 of the genome and
encode the nonstructural proteins. The multiple structural
proteins, including spike, nucleocapsid, envelope, and
membrane proteins are encoded by downstream open
reading frames (Fig. 2) [6–8]. SARS-CoV and MERS-CoV
belong to the beta coronavirus genus. However, SARS-
CoV belongs to lineage B, and MERS-CoV belongs to
lineage C along with bat coronaviruses HKU4 and HKU5.
As MERS-CoV and bat coronaviruses are part of lineage C
and MERS-CoV RNA was found in a bat sample in The
Kingdom of Saudi Arabia, researchers hypothesize that
bats may be a natural reservoir for MERS-CoV [9, 10].
Results from a recent study support that bats may be a
reservoir for MERS-CoV; however, camels and goats are
thought to be intermediate hosts [11]. In this study, MERS-
CoV was isolated from nasal secretions of MERS-CoV-
infected dromedary camels that had a short, mild disease
progression.
The suspected reservoir for SARS-CoV is the Chinese
horseshoe bat [2]. However, the mechanism of emergence
and adaptation to make the virus zoonotic is still not def-
initely understood [2]. SARS-CoV-like isolates from these
bats have up to 95% sequence similarity to human and
civet SARS-CoV. During the initial outbreak, SARS-CoV
was originally isolated from palm civets found in a Chinese
market; but, SARS-CoV was not found in the wild palm
civet population [12]. Bats harbor many coronaviruses and
are considered the main reservoir for later infections in an
intermediate host, such as civets or camels, which spread
the disease to humans [2]. Human-to-human transmission
has been most commonly associated with health-care
workers and those with close, unprotected contact with
infected patients [13, 14].
1.2 Clinical Features
The clinical features of MERS and SARS are similar and
can range from asymptomatic or mild disease to severe
pneumonia with acute respiratory distress syndrome
(ARDS) and multi-organ failure [15]. Although MERS and
SARS are clinically similar, the MERS mortality rate is
40% and SARS’s mortality rate is 10% [16]. Approxi-
mately 75% of MERS cases were associated with under-
lying comorbidities with a 60% mortality rate in this
subgroup (including cardiopulmonary abnormalities, obe-
sity, and diabetes). In contrast, 10–30% of patients with
SARS have comorbidities with a mortality rate of 46%
within this subgroup [15, 16].
The development of symptomatic MERS and SARS
mostly occurs in adults (median age of 50 years; 40 years
for SARS). MERS and SARS symptoms typically follow a
mean incubation time of * 5 days (range 2–13 and
2–14 days, respectively) and include fever, chills, cough
(some associated with blood), shortness of breath, myalgia,
headache, nausea, vomiting, diarrhea, sore throat, and
malaise [15–17]. Progression from mild to severe disease is
more rapid with MERS as compared to SARS with means
of 7 and 11 days, respectively [15]. Secondary bacterial
infections have occurred in patients with severe MERS;
however, the role of these coinfections in MERS patho-
genesis has yet to be determined [18–20]. Laboratory
abnormalities associated with MERS and SARS patients
include elevated lactate dehydrogenase, elevated liver
enzymes; thrombocytopenia; lymphopenia and leukopenia
[21–23].
1936 J. Dyall et al.
Radiographic abnormalities consistent with viral pneu-
monitis and ARDS are common in MERS and SARS.
Radiographic progression in the lower lobes has been
reported to be more rapid for MERS than SARS [21–23].
For SARS, disease in the lower lobes mimics pneumonia,
radiographic progression includes ground-glass opacifica-
tion and lobe thickening [17]. MERS-CoV (intact virus or
viral genome) is found at higher concentrations in the
lower respiratory tract than in the upper respiratory tract in
MERS patients and this may account for inefficient inter-
human transmission [15, 24]. Currently, no approved
therapeutics for patients with MERS or SARS are avail-
able, and clinical management has relied primarily on
supportive care.
Fig. 1 Maps of the severe acute respiratory syndrome (SARS) (a) and Middle East respiratory syndrome (MERS) (b) outbreaks with confirmed
case numbers
Therapeutics for MERS-CoV and SARS-CoV 1937
2 Therapeutic Agents
2.1 Clinical Usage
2.1.1 Treatment of SARS
Effectiveness of antiviral treatments used during the SARS
epidemic has been mainly based on case studies and ret-
rospective analysis of patient cohorts. Few randomized,
blinded, clinical trials of anti-SARS treatments were per-
formed, which adds complexity when interpreting the
available data (Table 1). Ribavirin, a nucleoside analog
that prevents RNA and DNA virus replication, was initially
used in the treatment of SARS due to its broad-spectrum
efficacy. For example, in a Taiwanese study, 51 SARS
patients were treated daily with fluoroquinolone antibiotics,
[levofloxacin (500 mg) or moxifloxacin (400 mg)] fol-
lowing diagnosis. Out of 51 patients, 44 SARS patients
were also treated intravenously (IV) with 2000 mg of rib-
avirin then orally daily with 1200 mg while 7 SARS
patients did not receive ribavirin. Corticosteroids, IV
methylprednisolone, or oral prednisolone were adminis-
tered as needed to treat worsening lung infiltrates and fever
[25]. Ribavirin treatment led to hypoxia and anemia and
increased risk for death in SARS patients. In a retrospective
analysis, a cohort of 229 patients from Hong Kong, Sin-
gapore, and Toronto were treated with ribavirin in con-
junction with corticosteroids, immunoglobulins, and/or
antibiotics [26]; ribavirin did not demonstrate efficacy.
Patients in Hong Kong and Singapore were treated with
ribavirin at 1200 mg orally at diagnosis, followed by oral
treatments with 2400 mg daily, or continual IV ribavirin
therapy [8 mg/kg every 8 h (h)]. In Toronto, patients
received ribavirin IV treatment with 2000 mg, followed by
1000 mg every 6 h for 4 days, and 300 mg every 8 h for
3 days. Unfortunately, fatality rates were similar between
the ribavirin-treated and control groups. Later, researchers
demonstrated that the ribavirin dosage required to be
effective against SARS-CoV in vitro was not clinically
achievable [27]. Ribavirin treatment also resulted in
adverse effects including anemia, hypoxemia and
decreased hemoglobin levels, and did not improve patient
outcome [26]. Due to the increasing adverse effects and
lack of efficacy, Health Canada stopped permitting the use
of ribavirin for SARS [25].
Table 1 Drug regimens used in the treatment of SARS
Treatment plan Treatment outcome
Ribavirin (oral/IV)
Antibiotics
± corticosteroids
± immunoglobulin
No increased positive outcome with ribavirin
compared to controls [25, 26]
Increased risk of anemia, hypomagnesemia,
hypoxia, or bradycardia with ribavirin
compared to ribavirin-naive patients
[25, 246]
Ribavirin (oral/IV)
Lopinavir/ritonavir
± corticosteroids
Fatality or acute respiratory distress syndrome
(ARDS) was reduced significantly from 28.8
to 2.4% [27]
IFN-alfacon-1
± corticosteroids
± antibiotics
Increased oxygen saturation
Increased clearance of lung abnormalities
Slight increase in creatinine kinase
concentrations [29, 247]
Fluoroquinolone
(IV)
Azithromycin (IV)
IFN-a (IM)
± corticosteroids
± Immunoglobulins
± thymic peptides/
proteins
No increased positive outcome [248]
Quinolone (IV)
Azithromycin (IV)
± IFN-a
± corticosteroids
No increased positive outcome [248]
Levofloxacin
Azithromycin
± IFN-a
± corticosteroids
Increased survival
Increased clearance of lung abnormalities
[248]
IFN interferon, IM intramuscular, IV intravenous, SARS severe acute
respiratory syndrome
Fig. 2 Genomes of Middle East respiratory syndrome coronavirus
(MERS-CoV) and severe acute respiratory syndrome coronavirus
(SARS-CoV) indicating the open reading frames for nonstructural (1a
and 1b) and structural proteins (numbered 3–9, and E, M, N, S).
E envelope, M membrane, N nucleocapsid, S Spike
1938 J. Dyall et al.
Additional studies tested the efficacy of ribavirin in
conjunction with lopinavir, an anti-retroviral agent. Lopi-
navir demonstrated in vitro activity against SARS-CoV
[28]. In a non-randomized, open-enrollment trial of 152
suspected SARS patients [27], all patients were treated
with ribavirin and corticosteroids similar to the previously
described studies. In addition, 41 of the confirmed SARS
patients were also treated with a combination of lopinavir
(400 mg) and ritonavir (100 mg). Mean viral loads in
nasopharyngeal swabs within this treatment group
decreased to undetectable levels by day 10. Overall, SARS-
related symptoms subsided, disease progression was
milder, and no adverse effects were reported as compared
to the historical control group.
In an open-label, non-randomized study of 22 SARS
patients, 9 patients who received subcutaneous (SC)
injections of interferon (IFN)a, alfacon-1, for 10 days at an
initial dose of 9 lg/day for 2 days increasing to 15 lg/daywith disease progression. All 9 patients survived with
minor adverse effects [29].
2.1.2 Treatment of MERS
The evaluation of treatments in MERS patients has been
hampered as high-quality clinical data from randomized
clinical trials are limited. Ribavirin (with or without IFN,
or corticosteroids) was the primary treatment during the
MERS outbreak. In a retrospective analysis, a cohort of 20
patients was treated with oral ribavirin and SC pegylated
IFN-a2a at a dose of 180 lg/week for 2 weeks (Table 2)
[30]. The initial dose of ribavirin was 2000 mg, followed
by a 200–1200 mg dose depending on creatinine clearance.
A group of 24 patients that received supportive care and
corticosteroids were considered the control group. At
14 days after confirmed diagnosis of MERS, survival was
increased in the treated group (70%) compared to the
control group (29%). By 28 days post-diagnosis, 30% of
treated subjects survived versus 17% of the control group
[30]. In an additional case study, a 69-year-old Greek
patient who contracted MERS in Jeddah was treated with
oral lopinavir/ritonavir (400/100 mg twice daily), pegy-
lated IFN (180 lg SC once per week for 12 weeks), and
ribavirin (2000 mg initial dose; 1200 mg every 8 h for
8 days, initiated on day 13 post-diagnosis). Two days after
treatment initiation, viremia could not be detected; how-
ever, viral RNA was detected in several patient samples
(feces, respiratory secretions, and serum) up to 14 weeks
post-diagnosis. Despite prolonged survival, the patient
succumbed from septic shock 2 months post-diagnosis
[31]. An ongoing randomized clinical trial in Saudi Arabia
is evaluating treatment of MERS patients with IFN-b1b in
combination with lopinavir/ritonavir [32].
2.2 Drugs with Repurposing Potential
for Treatment of Coronaviral Infections
Drug repurposing is an attractive alternative drug discovery
strategy because it eliminates many steps usually required
at the early phase of drug development. Over the past
decade, interest in drug repurposing has increased as
pharmaceutical companies are challenged with decreasing
product pipelines, high costs associated with de novo drug
discovery, and the imminent expiration of many drug
patents. Some examples for successfully repurposed drugs
include Viagra (Pfizer) for erectile dysfunction (original
indication: angina) and raloxifene (Eli Lilly) for treatment
of invasive breast cancer (original indication:
osteoporosis).
The time required for traditional drug development is
often discordant with the urgent need for novel therapies
for emerging infectious diseases such as SARS and MERS.
Outbreaks can occur anywhere in the world and frequently
in resource-limited settings. Commonly, the treatment
strategies that are available for emerging infectious dis-
eases are less than adequate to improve patient outcome.
Although specific antivirals for MERS-CoV and SARS-
CoV are in development, drug repurposing could present
an important arm in generating additional therapeutics for
future coronaviruses. First, if these drugs are confirmed to
have beneficial effects in vitro and in animal studies, they
could be used to build a panel of approved drugs for use as
a first-line of defense for newly emerging coronaviruses.
Second, these drugs could be made accessible relatively
quickly to patients under Emergency Use Authorization.
Extending the choices of treatment by generating a panel of
Table 2 Drug regimens used in the treatment of MERS
Treatment plan Treatment outcome
Ribavirin (oral/
IV)
IFN-a2b
Corticosteroids
Late treatment administration. Disease
progression delayed—all patients died [249]
Ribavirin (oral/
IV)
PEGylated IFN-
a2a (IV)
± corticosteroids
Treatment initiated 0–8 days after diagnosis
Adverse effects: significant decreases in
hemoglobin and absolute neutrophil count
(baseline count lower in treatment group) [30]
Ribavirin (oral/
IV)
Lopinavir/
ritonavir
IFN-a2b
No detectable viral RNA in serum after 2 days of
therapy
Adverse effects: ribavirin discontinued due to
jaundice, hyperbilirubinemia
Died of septic shock 2 months, 19 days after
diagnosis [31]
IFN interferon, IV intravenous, MERS Middle East respiratory
syndrome
Therapeutics for MERS-CoV and SARS-CoV 1939
broad-spectrum antivirals would provide a real improve-
ment to healthcare communities struggling to cope during
an outbreak of emerging infections. A great example of
how repurposing can benefit in the search of treatments for
emerging infections is the drug zidovudine. Zidovudine
was originally developed in 1964 as a cancer drug. In 1985,
zidovudine was found to be active against human
immunodeficiency virus (HIV), and 2 years later it became
the first drug to be approved for the treatment of acquired
immunodeficiency syndrome (AIDS) [33].
A number of research groups have identified and
investigated the usefulness of approved drugs for the
treatment of viral infections including coronaviruses.
Below, we summarize several drug classes with antiviral
activity against MERS-CoV and SARS-CoV that have
repurposing potential (Fig. 3, Table 3). Some of the drugs
described have activity against other virus families indi-
cating potential broad-spectrum applications and have
already been in clinical use for treating other viral infec-
tions. We would like to emphasize that none of the thera-
peutics described in this section are recommended for
clinical use outside a clinical trial setting.
2.2.1 Antidiarrheal Agents
Loperamide, an approved anti-diarrheal agent, is on the
World Health Organization (WHO) Model List of Essential
Medicines and is available in many countries. The drug
acts on the opioid receptor and reduces intestinal motility
[34]. Results from pharmacokinetic (PK) studies show that
oral loperamide is well absorbed from the gut with less
than 1% of the drug entering systemic circulation [35].
Loperamide demonstrated anti-MERS-CoV, anti-SARS,
and anti-HCoV229E activity in an in vitro screen of
approved drugs [36], although the mechanism of action is
unknown. Interestingly, loperamide was suggested for
limiting gastrointestinal fluid and electrolyte losses in
patients with Ebola virus disease (EVD) [37].
2.2.2 Antimalarial Agents
The antimalarial agents, chloroquine (CQ), amodiaquine,
and mefloquine have activity against SARS-CoV and
MERS-CoV in vitro [36, 38, 39]. CQ is a U.S. Food and
Drug Administration (FDA)-approved antimalarial agent
that is also used to treat autoimmune disease such as
rheumatoid arthritis due to its anti-inflammatory effects
[40]. CQ has activity against a number of viruses in vitro
and in vivo including flaviviruses [dengue virus (DENV)],
Togaviruses [chikungunya virus (CHIKV)], paramyx-
oviruses (Hendra, Nipah virus), influenza viruses, HIV,
and filoviruses [Ebola virus (EBOV)] [41–47].
Several mechanisms of action have been identified for
the antiviral effect of CQ and suggest that the drug acts
nonspecifically at virus entry or at the later stages of virus
production. CQ accumulates within acidic organelles such
as endosomes, Golgi vesicles, and lysosomes, where the
drug is protonated resulting in increased pH within the
vesicle [48]. Viruses depend on these acidic organelles for
entry, viral replication, and maturation of virus progeny.
Similarly, MERS-CoV entry into cells depends on several
proteases. Dipeptidyl peptidase 4 (DPP4) acts as functional
virus receptor [49], and cellular proteases [e.g. type II
transmembrane serine protease (TMPRSS2) and members
of the cathepsin family] activate the viral spike (S) glyco-
protein [50]. CQ may have an effect on any of these pro-
teases. CQ also affects the glycosylation step within the
Golgi that directs trafficking and maturation of viral pro-
teins [51–53]. For SARS-CoV, the antiviral activity of CQ
has also been attributed to a deficit in glycosylation of the
receptor angiotensin-converting enzyme 2 (ACE2) [54].
The broad-spectrum antiviral activity makes CQ an
attractive antiviral for repurposing and treating coronaviral
and other emerging viral infections. In vivo activity of CQ
in MERS or SARS animal models has not yet been
reported. However, the antiviral activity of the drug has
been evaluated against other viruses in preclinical and
clinical studies with mixed results. CQ plasma steady state
concentrations in mice are similar to those reported for
humans (10 lM) and are within range of the EC50 values
determined for MERS-CoV (3.6 lM) and SARS-CoV
(2.3 lM) [43, 55]. Preclinical studies with CQ in mice
against other viruses have shown survival benefits for
influenza and EBOV infections.
In clinical studies, CQ was effective at reducing viral
loads in asymptomatic HIV patients [56, 57], but results of
CQ treatment of CHIKV and DENV infections were mixed
[58, 59].
In summary, CQ has broad-spectrum potential and the
information gained from studies on other viruses can be
used to plan the most appropriate strategies for evaluating
its specific clinical value for treating for MERS-CoV and
SARS-CoV infections. CQ has several advantages includ-
ing rapid absorption from gastrointestinal tract, low cost,
cFig. 3 Candidate drugs for repurposing for coronaviral infections.
Several drug classes (A through I) have been studied, and the steps/
processes of the viral replication cycle that they most likely target are
indicated. AKT serine/threonine kinase, CAD cationic amphiphilic
drug, Cyps cytochrome P-450s, E envelope, ER endoplasmic retic-
ulum, ERGIC ER–Golgi intermediate compartment, ERK extracellu-
lar signal-reduction kinase, IFN interferon, MAPK mitogen-activated
protein kinase, M membrane, MPA mycophenolic acid, mTOR
mechanistic target of rapamycin, N nucleocapsid, NFAT nuclear
factor of activated T cells, ORF open reading frame, PI3K phospho-
inositide 3-kinase, S Spike
1940 J. Dyall et al.
Therapeutics for MERS-CoV and SARS-CoV 1941
and very effective biodistribution. CQ may be an excellent
candidate for combinatorial treatments with other antivi-
rals. However, considerable challenges remain for the
treatment of viral infections including increased under-
standing of the pharmacodynamics of CQ, achievement of
sufficient plasma concentrations in patients, and toxicity
concerns [60]. Importantly, hydroxychloroquine, a CQ
derivate, may provide an alternative due to lower toxicity
and similar pharmacology profile [55].
A related antimalarial drug, amodiaquine, also has
activity in vitro against MERS-CoV and SARS-CoV [38].
Previous investigations have demonstrated that amodi-
aquine inhibits filovirus replication, and the mechanism of
action is hypothesized to be similar to that of CQ [43].
Amodiaquine is well tolerated and is commonly used for
malaria treatments in many countries. Further, amodi-
aquine in combination with artesunate was administered to
EVD patients during the 2013–2016 epidemic, and the
resulting decrease in fatality rates may have been associ-
ated with the use of amodiaquine as an antimalarial agent
[61]. Nonhuman primate (NHP) studies are currently
underway to investigate the effect of amodiaquine treat-
ment on EVD [62].
Mefloquine, a synthetic analog of quinine, is another
antimalarial drug with activity against MERS-CoV and
SARS-CoV [38]. It belongs to the WHO Model List of
Essential Medicines. Mefloquine is known to penetrate the
blood-brain barrier and was found to inhibit JC virus
infection and replication at concentrations generally
achieved in the brains of patients given mefloquine for
malaria [63] leading to the clinical evaluation of this drug
for the treatment of progressive multifocal leukoen-
cephalopathy [64, 65]. In 2013, the FDA added a boxed
warning to the US label of mefloquine regarding the
potential for neuropsychiatric side effects. Additional
investigations are warranted to determine if amodiaquine
or mefloquine have value for repurposing for treatment of
MERS or SARS.
2.2.3 Cyclophilin Inhibitors
Cyclophilins are ubiquitous host proteins believed to have
multiple roles in trafficking, protein folding and T cell
activation [66]. Cyclosporine A (CysA), forms a complex
with cyclophilin A, thereby blocking T cell activation.
CysA is licensed for use in organ transplantation to sup-
press the immune response. CysA has also been shown to
inhibit coronaviruses including SARS-CoV and MERS-
CoV effectively in cell culture [67, 68]; however, the
mechanism has yet to be determined. There is increasing
evidence that cyclophilins are involved in viral replication
of RNA viruses such as hepatitis C virus (HCV) and West
Nile virus, and this may also apply to coronaviruses [69].
Although the immunosuppressive properties of CysA are
considered a risk for treating viral infections in patients,
nonimmunosuppressive analogs of CysA that bind to
cyclophilins with higher affinity have been developed and
some are in clinical trials as HCV therapeutics [70–72].
2.2.4 Interferons
Interferons (IFNs) are approved by the FDA for other
indications such as hepatitis C. Although IFN-a reduced
SARS-CoV replication in mice and NHPs [73, 74], efficacy
of IFN-a treatment in SARS patients was mixed (see
Sect. 2.1.1). From in vitro studies, another type I inter-
feron, IFN-b1a, may be more effective than IFN-a either
alone or in combination with IFN-c [75–77]. Combinations
of IFN-b and -c were synergistic against SARS-CoV
in vitro [77].
With regards to MERS, in vitro and in vivo preclinical
studies have indicated that IFN-a2b alone or in combina-
tion with ribavirin, may have a therapeutic effect if given
early in disease [78, 79]. In clinical trials, however, IFN-
a2b (given in combination with other treatments) did not
lead to a significant benefit to patients (see Sect. 2.1.2).
IFN-b1a (EC50 = 1.37 IU/mL) was superior in activity
against MERS-CoV infection in vitro compared to IFN-
a2a, IFN-a2b, and IFN-c; these IFNs had EC50 values of
160.8, 21.4, and 56.5 IU/mL, respectively [80]. IFN-b1b is
currently under evaluation for MERS-CoV in a randomized
clinical trial (in combination with lopinavir/ritonavir) [32].
Investigating the IFN-b subtypes (1a and 1b) in combina-
tion with other antivirals may be worthwhile as potential
synergistic combinations could reduce the effective drug
dosage and IFN-associated adverse effects.
2.2.5 Kinase Inhibitors
Many cellular processes are regulated independently of
changes in transcription or translation through kinase-me-
diated cell signaling pathways. As a testament to the bio-
logical importance of kinases, there have been over 500
kinases identified along with more than 900 genes encod-
ing proteins with kinase activity [81, 82]. As of April 2015,
28 kinase inhibitors have been granted approval by the US
FDA with over half gaining approval from 2012–2015.
Further, kinases are the most frequently targeted gene class
in cancer therapy, second only to the G protein-coupled
receptors as therapeutic targets [83, 84].
The therapeutic potential for host-targeted
immunomodulatory agents in viral infections has received
considerable attention [85–87]. Recently, Dyall et al.
identified two Abelson (Abl) kinase inhibitors (imatinib
and dasatinib) that inhibited MERS-CoV and SARS-CoV
infection through a cell-screening assay [38]. Both
1942 J. Dyall et al.
Table
3Clinically
developed
drugswithactivityagainst
MERS-CoV
andSARS-CoV
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
Antidiarrhealagents
Loperam
ide
hydrochloride
MERS-CoV:
4.8
lM[36]
SARS-CoV:
5.9
lM
HCoV-229E-G
FP:
4lM
[36]
Approved
fortreatm
entofdiarrhea
Welltolerated,
commonly
used
WHO
list
ofessential
medicines
Antimalaria
agents
Amodiaquine
hydrochloride
MERS-CoV:
6.2
lM[38]
SARS-CoV:
1.3
lM[38]
EBOV
[43];MARV
[43];DENV
[250]
Approved
fortreatm
entofmalaria
Welltolerated,
commonly
used
Chloroquine
diphosphate
MERS-CoV:
3–6.3
lM
[36,38]
SARS-CoV:
4.1–8.8
lM
[36,38,39]
Invitro:
CHIK
V[251,252];DENV
[42,47];
HeV
,NiV
[46];HIV
-1[41];
FLUAV
[44];EBOV,MARV
[43];
SFV,SIN
V[45]
Invivo:
Mixed
dataonefficacy
against
EBOV
inmice[43,60];noefficacy
against
EBOV
inham
sters,guinea
pigs[60];FLUAV:70%
survivalin
mice[253]
Approved
fortreatm
entofmalaria
Clinical
trials
fortreatm
entofCHIK
V,
DENV,FLUAV,andFLUBV
infectionsshowed
noim
pacton
disease
[58,59,254]
Welltolerated,
commonly
use
WHO
list
ofessential
medicines;
WHO
listofpotential
EVD
treatm
ents;dosing,form
ulation
needto
beoptimized
fortreatm
ent
ofviral
infections[54]
Hydroxychloroquine
sulfate
MERS-CoV:
8.3
lM[38]
SARS-CoV:
8.0
lM[38]
Invitro:
DENV
[42];HIV
-1[255];JC
V[63]
Approved
fortreatm
entofmalaria
Welltolerated,
commonly
used
WHO
list
ofessential
medicines
Mefloquine
MERS-CoV:
7.4
lM[38]
SARS-CoV:
15.6
lM[38]
Approved
fortreatm
entofmalaria
Clinical
trials
fortreatm
entofJC
V
infection:mixed
results[64,65]
Black
boxwarning:
potential
neuropsychiatric
sideeffects
WHO
list
ofessential
medicines
Cyclophilin
inhibitors
CyclosporinA
MERS-CoV
[67]
SARS-CoV
[68]
HCV
[256];WNV
[257];JEV
[258]
VSV
[259];HIV
-1[260]
Approved
forim
munosuppression
duringorgan
transplantation
Immunosuppression
undesirable
for
infectiousdiseases
WHOlistofessentialmedicines;non-
immunosuppressiveanalogsare
available
[70–72]
Interferons
IFN-a2a
MERS-CoV:
160.8
U/m
L[80]
Approved
fortreatm
entofhepatitis
B
andC
Welltolerated
Therapeutics for MERS-CoV and SARS-CoV 1943
Table
3continued
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
IFN-a2b
MERS-CoV:
21.4
U/m
L[80]
SARS-CoV:6500-
4950U/m
L[75]
Approved
fortreatm
entofmelanoma
IFN-b1a,
1b
MERS-CoV:
1.4
U/m
L[80]
SARS-CoV:
95–105U/m
L
[75]
Approved
fortreatm
entofmultiple
sclerosis
IFN-c
MERS-CoV:
56.5
U/m
L[80]
SARS-CoV:
1700–2500U/
mL[115]
Approved
fortreatm
entofchronic
granulomatousdisease
Kinaseinhibitors
Dabrafenib
MERS-CoV:45%
inhibitionat
10lM
[89]
Approved
fortreatm
entofcancers
Welltolerated
Dasatinib
MERS-CoV:
5.5
lM[38]
SARS-CoV:
2.1
lM[38]
Invitro:
BKPyV
[261];EBOV
[100];HIV
-1
[262];DENV
[263]
Invivo:
VACV:noactivityin
mice[264]
Approved
fortreatm
entofcancers
Welltolerated
ABL1inhibitor,CAD
Immunosuppressiveeffectsin
vivo
may
precludeuse
asanti-infective
Everolimus
MERS-CoV:56%
inhibitionat
10lM
[89]
DENV,CPXV,RSV,FLUAV
[265]
Approved
immunosuppressantfor
cancertreatm
entandpreventionof
organ
rejection;reducedincidence
of
HHV-5
incardiacandrenaltransplant
patients
[266]
Welltolerated
mTOR
inhibitor[267]
Imatinib
mesylate
MERS-CoV:
17.7
lM[38]
SARS-CoV:
9.8
l M[38]
Invitro:
BKPyV
[261];HHV
[268];HCV
[149];MPXV,VACV,VARV[264]
Invivo:
VACV:100%
survival
inmice[264]
Approved
fortreatm
entofcancers
Welltolerated
ABL1inhibitor
Miltefosine
MERS-CoV:28%
at10lM
[89]
HSV-2
[269]
Antimicrobialapproved
fortreatm
entof
leishmaniasis;investigational
drugfor
treatm
entofam
oebainfections
Welltolerated
Aktinhibitor
WHO
list
ofessential
medicines
1944 J. Dyall et al.
Table
3continued
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
Nilotinib
MERS-CoV:
5.5
lM[38]
SARS-CoV:
2.1
lM[38]
Selumetinib
sulfate
MERS-CoV:
C95%
inhibition
at10lM
[89]
InPhaseII/IIItrialsforcancertreatm
ent
Welltolerated
MEK
inhibitor
Sirolimus
MERS-CoV:61%
inhibitionat
10lM
[89]
HIV
-1[270];HHV-5
[271]
Approved
immunosuppressantin
transplant;reducedviral
replicationin
HIV
-orHCV-positivetransplant
patients
[272–274]
Welltolerated
mTOR
inhibitor
Sorafenib
MERS-CoV:93%
inhibitionat
10lM
[89]
BKPyV
[261];HHV
[275];EV71
[276];RVFV
[277]
Approved
fortreatm
entofcancers
Welltolerated
Trametinib
MERS-CoV:
C95%
inhibition
at0.1
lM[89]
Approved
fortreatm
entofcancers
Welltolerated
Wortmannin
MERS-CoV:40%
inhibitionat
10lM
[89]
HSV-2
[269];HIV
-1[278]
Indevelopmentfortreatm
entofcancer
Toxicityissues.
Derivatives
arein
PhaseIclinical
trials
Neurotransm
itterinhibitors
Astem
izole
MERS-CoV:
4.9
lM[38]
SARS-CoV:
5.6
lM[38]
Previouslyapproved
antihistamine
Withdrawnin
1999
because
ofrare
arrhythmias
Benztropine
mesylate
MERS-CoV:
16.6
lM[38]
SARS-CoV:
21.6
lM[38]
HCV
[102],EBOV,MARV
[100,101,104]
Approved
anticholinergic
fortreatm
ent
ofParkinson’s
disease
Welltolerated
Chlorphenoxam
ine
MERS-CoV:
12.7
lM[38]
SARS-CoV:
20.0
lM[38]
Approved
antihistamineand
anticholinergic
fortreatm
entof
Parkinson’s
Disease
Welltolerated
Chlorpromazine
hydrochloride
MERS-CoV:
4.9–9.5
lM
[36,38]
SARS-CoV:
8.8–13.0
lM
[36,38]
EBOV
[96];JU
NV
[94];SV40[279]
Approved
fortreatm
entof
schizophrenia
Welltolerated
CAD
Therapeutics for MERS-CoV and SARS-CoV 1945
Table
3continued
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
Clomipramine
hydrochloride
MERS-CoV:
9.3
lM[38];
SARS-CoV:
13.2
lM[38]
EBOV
[100]
Approved
fortreatm
entofdepression
Welltolerated
WHO
list
ofessential
medicines
Fluphenazine
hydrochloride
MERS-CoV:
5.9
lM[38];
SARS-CoV:
21.4
lM[38]
HCV
[149];EBOV
[100]
Approved
fortreatm
entofchronic
psychoses
Welltolerated
WHO
list
ofessential
medicines;
CAD
Fluspirilene
MERS-CoV:
7.5
lM[38]
SARS-CoV:
5.9
lM[38]
Approved
fortreatm
entof
schizophrenia
Welltolerated
Promethazine
hydrochloride
MERS-CoV:
11.8
lM[38]
SARS-CoV:
7.5
lM[38]
Antipsychoticapproved
forsedation
Welltolerated
Thiothixene
MERS-CoV:
9.3
lM[38]
SARS-CoV:
5.3
lM[38]
Antipsychoticapproved
fortreatm
entof
schizophrenia
Welltolerated
Triethylperazine
maleate
MERS-CoV:
7.8
lM[38]
SFV
[45],CHIK
V[45,99]
Approved
antiem
etic
Welltolerated
Triflupromazine
hydrochloride
MERS-CoV:
5.8
lM[38];
SARS-CoV:
6.4
lM[38]
Approved
antipsychotic
Serioussideeffects
includeakathisia.
CAD
Nucleicacid
synthesis
inhibitors
Gem
citabine
hydrochloride
MERS-CoV:
1.2
lM[38]
Invitro:
FLUAV
[125]
Invivo:
MiceMuLV
[124]
Approved
fortreatm
entofcancers
Welltolerated
WHO
list
ofessential
medicines
Mizoribine
SARS-CoV:3.5-
16lg
/mL[114]
or[
40lM
[111]
HCV
[123];BVDV
[280]
Approved
immunosuppressantin
organ
transplantationandrheumatic
diseases.
Welltolerated
Mycophenolicacid
MERS-CoV:
0.17lg
/mL
[112]or2.87lM
[80]SARS-CoV:
[30lM
[111]
DENV
[118];CHIK
V[252];FLUAV
[112];SFV,SIN
V[45]
Approved
immunosuppressantin
organ
transplantation
FDA
alert:risk
of
activationoflatent
herpes
infections
1946 J. Dyall et al.
Table
3continued
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
Proteaseinhibitors
Lopinavir
Invitro:
MERS-CoV:
8lM
[36]
SARS-CoV:
24.4
lM[36]
Invivo:
67%
survival
in
MERS-infected
NHPs[121]
HIV
-1[136]
HPV
[281]
Approved
fortreatm
entofHIV
infections
Clinical
trialfortopical
treatm
entof
cervical
cancer
Clinical
trialin
SARSpatients
[27]
Welltolerated
Lopinavir/ritonavir:WHO
listof
essential
medicines
E-64-D
MERS-CoV:
1.27lM
[38];
SARS-CoV:
0.76lM
[38]
EBOV
[134,135]
Cysteineproteaseinhibitor
PhaseIIIfortreatm
entofmuscular
dystrophy
Welltolerated
K1177
MERS-CoV,
SARS-CoV,
HCoV-229E
(VLP)[133]
EBOV,MARV,NiV
(VLP)[133]
Cysteineproteaseinhibitor
Inclinical
developmentfortreatm
entof
Chagas
disease
Welltolerated
Cam
ostat
mesylate
Invitro:
MERS-CoV
[132],SARS-
CoV
[132]
Invivo:
SARS:activityin
mice[133]
Invitro:
FLUAV,FLUBV
[130];no
inhibitionofEBOV
[133]
Cellularserineproteaseinhibitor
Inclinical
developmentforchronic
pancreatitis
Welltolerated
SARS-CoVspread
isdriven
byserine
proteaserather
than
cysteine
protease[133]
Protein
synthesis
inhibitors
Anisomycin
MERS-CoV:
0.003lM
[38]
SARS-CoV:
0.19lM
[38]
PV
[147];EMCV
[146]
Antibioticin
clinical
developmentfor
treatm
entofam
oebiasis
Emetine
hydrochloride
MERS-CoV:
0.014lM
[38]
SARS-CoV:
0.05lM
[38]
FLUAV
[112];EMCV
[146]
Approved
antibioticfortreatm
entof
amoebiasisin
somecountries
Sideeffectsinclude
nausea;
derivatives
withless
side
effectsavailable
Omacetaxine
mepesuccinate
MERS-CoV:
9.07lM
[38]
MHV:0.012lM
[139]
HBV
[148]
Approved
forchronicmyeloid
leukem
iaWelltolerated
Therapeutics for MERS-CoV and SARS-CoV 1947
Table
3continued
Drugclassmem
bers
Activityagainst
coronavirusesa
Activityagainstother
viruses
Clinical
status
Knownsafety
issues
Comments
Selectiveestrogen
response
modulator
Tam
oxifen
citrate
MERS-CoV:
10.1
lM[38];
SARS-CoV:
92.9
lM[38]
HCV
[282];HSV-1
[283]
Approved
fortreatm
entofbreastcancer
Black
boxwarning:
uterinecancer,
bloodclots,stroke
WHO
list
ofessential
medicines;
CAD
Toremifenecitrate
MERS-CoV:
12.9
lM;SARS-
CoV:12lM
[38]
Invitro:
HCV
[149];EBOV
[100,151];
SUDV;RAVV;MARV
[151]
Invivo:
EBOV:50%
survival
inmice[151]
Approved
fortreatm
entofbreast
cancer.
Black
boxwarning:
cardiaceffect
(QT
prolongation)in
patientswith
hypokalem
ia
WHO
list
ofpotential
EVD
treatm
ents;CAD
Sterolmetabolism
inhibitors
Terconazole
MERS-CoV:
12.2
lM[38];
SARS-CoV:
15.3
lM[38]
EBOV-V
LP[152];EVD
[100]
Approved
topical
antifungal
for
treatm
entofvaginal
yeast
infections
Would
requireIN
D
fororaluse
CAD
Triparanol
MERS-CoV
5.3
lM[38]
EBOV-V
LP[152]
Developed
forloweringserum
cholesterol;withdrawn
Acute
cataract
form
ation
CAD
ABL1abelsonmurineleukem
iaviral
oncogenehomolog1,Akt
protein
kinaseB,BKPyV
BK
polyomavirus,
BVDV
Bovineviral
diarrhea
virus,
CAD
cationic
amphiphilic
drug,CHIK
Chikungunyavirus,
CPXV
Cowpox
virus,
CVsCoxsackie
viruses,
DENV-1
Denguevirus-1,DENV-2
Denguevirus-2,EBOV
Ebola
virus,
EC50effectiveconcentration
50,EMCV
Encephalomyocarditisvirus,EV71Enterovirus71,FCoVFelinecoronavirus,FDAFoodandDrugAdministration,FLUAVInfluenza
Avirus,FLUBVInfluenza
Bvirus,HBVHepatitisBvirus,
HCV
hepatitis
Cvirus,
HeV
Hendra
virus,
HHV-1
Human
herpesvirus-1(herpes
simplexvirus-1),
HHV-2
human
herpresvirus-2(herpes
simplexvirus-2),
HHV-5
Human
herpesvirus-5
(cytomegalovirus),HIV-1
Human
immunodeficiency
virus-1,HIV-2
Human
immunodeficiency
virus-2,HPVHuman
papillomavirus,HRVsHuman
rhinoviruses,INDInvestigationalnew
drug,
JCVJohnCunningham
virus,JU
NVJunin
virus,MARVMarburg
virus,MEK
MAPK/ERK
kinase,
MERS-CoVMiddle
Eastrespiratory
syndromecoronavirus,MHVMouse
hepatitis
virus,
MPXVMonkeypoxvirus,mTORmechanistictargetofrapam
ycin,MuLVMurineleukem
iavirus,NiV
Nipah
virus,PVPoliovirus,RAVVRavnvirus;RSVRespiratory
syncytialvirus,RVFVRift
Valleyfever
virus,SARS-CoVSevereacuterespiratory
syndromecoronavirus,SFVSem
likiforestvirus,SFTSVSeverefever
withthrombocytopeniasyndromevirus,SINVSindbisvirus,SUDV
Sudan
virus,SV40Sim
ianvirus40;VACVVaccinia
virus,VARVVariola
virus,VLPvirus-likeparticles,WHO
WorldHealthOrganization
aAntiviral
activityisexpressed
interm
sofEC50unless
otherwisenoted
1948 J. Dyall et al.
compounds significantly inhibited MERS-CoV and SARS-
CoV with micromolar EC50 values and low cytotoxicity.
Abl2 has been identified as critical for MERS-CoV and
SARS-CoV virus entry, and may be the target that imatinib
inhibits to block entry of both viruses [88]. A recent sys-
tems kinome analysis investigation of in vitro MERS-CoV
infection suggested that ERK/MAPK and PI3 K/Akt/
mTOR signaling pathways were specifically modulated
during infection [89]. Subsequent analysis of licensed
kinase inhibitors targeting these pathways demonstrated
that kinase inhibitors targeting the ERK/MAPK signal
pathway (selumetinib and trametinib) inhibited MERS-
CoV infection by C 95% when added pre- or post-infection
[89]. Further, trametinib demonstrated significantly stron-
ger inhibitory activity against MERS-CoV than selume-
tinib suggesting that specific intermediates of the ERK/
MAPK signaling pathway may represent crucial foci dur-
ing early (viral entry) and late (viral replication) events in
the viral life cycle. In contrast, sorafenib, an inhibitor of
Raf-1 and B-Raf, components of the ERK/MAPK signaling
pathway, and vascular endothelial growth factor receptor 2
(VEGFR2), inhibited MERS-CoV infection by [90%
when added to cells prior to infection; however, the inhi-
bitory activity was reduced to \30% when added post-
infection suggesting Raf kinases were primarily involved
in early viral life cycle events. In addition, the inhibitory
activity dabrafenib, a Raf kinase inhibitor, was also largely
ablated when added post-infection. Miltefosine, an alkyl
phospholipid, considered to be an inhibitor of protein
kinase B (Akt), garnered FDA approval for infectious
disease-related treatments (cutaneous or mucosal leishma-
niasis) [90]. In 2013, miltefosine became directly avail-
able from the US Centers for Diseases Control and
Prevention for the treatment of free-living amoeba infec-
tions [91]. Pre-treatment of cells with miltefosine reduced
MERS-CoV infection by 28%, but had no effect when
added post-infection [89]. In contrast, inhibition of mTOR
with sirolimus or everolimus reduced MERS-CoV infec-
tion by * 60% when added prior pre- or post-infection
suggesting a critical role for mTOR in MERS-CoV infec-
tion. A recent clinical investigation by Wang et al. [92]
evaluated sirolimus and corticosteroids in addition to
standard antiviral treatment in a randomized controlled trial
in patients with severe H1N1 pneumonia and acute respi-
ratory failure [92]. Importantly, the addition of sirolimus
was associated with improved patient outcomes including
decreased hypoxia and multi-organ dysfunction, reduced
mean times for liberation from mechanical ventilation, and
increased clearance of virus. Thus, it may be prudent to
extend the study of repurposed kinase inhibitors beyond
stand-alone therapeutic investigations and also consider
their potential as adjunctive therapies.
2.2.6 Neurotransmitter Inhibitors
Numerous neurotransmitter receptors inhibitors showed
activity against MERS-CoV and SARS-CoV infection [38].
These drugs were initially developed as antipsychotics,
antihistamines, and sedatives. Five neurotransmitter
receptor antagonists belong to the chemical class of phe-
nothiazines: chlorpromazine, triflupromazine, thiethylper-
azine, promethazine and fluphenazine. Phenothiazines were
breakthrough medications developed in the 1950s for
treating mental health patients and reduced episodes of
bizarre behavior, hallucinations, and irrational thoughts
[93]. Although phenothiazines primarily block dopamine
receptors, they also have anticholinergic, antihistamine,
and antiemetic effects.
The phenothiazines, chlorpromazine and triflupro-
mazine, are approved antipsychotics. Chlorpromazine has
been used off-label for short-term treatment of nausea and
migraines. Triflupromazine is used to treat severe emesis,
but the drug has more serious side effects than chlorpro-
mazine including akathisia and tardive dyskinesia. The
antiviral effect of chlorpromazine has been extensively
studied, and the drug interferes with clathrin-mediated
endocytosis, a process that many viruses exploit for host
cell entry. Chlorpromazine inhibits entry of Junin virus
[94], West Nile virus [95], EBOV [96], HCV [97], and
Japanese encephalitis virus [98] suggesting broad-spectrum
activity that could be exploited early during a novel virus
outbreak. Chlorpromazine may have similar effects on
coronaviruses as the drug effectively inhibits MERS-CoV,
SARS-CoV, and human coronavirus 229E expressing
green fluorescent protein [36, 38]. However, time-of-ad-
dition studies indicate that the inhibitory activity against
MERS-CoV is retained whether added pre- or post-infec-
tion suggesting that there are additional effects to clathrin-
mediated entry impairment [36].
Thiethylperazine is an approved antiemetic. Both
chlorpromazine and thiethylperazine have been shown to
inhibit alphaviruses, Semliki forest virus (SFV) and
chikungunya virus (CHIKV) [45, 99]. As these drugs cross
the brain-blood barrier, use of these drugs could be bene-
ficial in the treatment of CHIKV, including common neu-
rologic complications. Promethazine is an antihistamine
used as a sedative in many countries under different brand
names, but also acts as a weak anti-psychotic activity.
Fluphenazine is a common antipsychotic used to treat
chronic psychoses (primarily schizophrenia) and belongs to
the WHOModel List of Essential Medicines. Promethazine
and fluphenazine have shown in vitro activity against
MERS-CoV and SARS-CoV and may have value as can-
didates for repurposing for coronaviral infections [38].
Benztropine mesylate, an approved anticholinergic used
to treat Parkinson’s, had activity against MERS-CoV and
Therapeutics for MERS-CoV and SARS-CoV 1949
SARS-CoV [38]. Benztropine was also identified in other
screens of clinically approved drugs for antiviral activities
against HCV and EBOV [100–102]. Although the detailed
mechanism of action is unknown, HCV studies indicate
that benztropine inhibits at a virus entry step, while not
interfering with viral genome replication, transcription or
production of viral progeny or virus production of viral
progeny [102]. It has been argued that a virus entry inhi-
bitor may have value in decreasing the incidence of relapse
in chronic HCV patients that receive liver transplants
[102]. However, the peak plasma concentrations of ben-
ztropine may be too low to be effective for treating an
acute infection [103]. Benztropine was also independently
identified in two drug screens for EBOV antivirals
[100, 101]. A recent report suggests that a step after virus
attachment, but prior to viral/cell membrane fusion is tar-
geted by benztropine [104].
Clomipramine, a tricyclic antidepressant, and thiothix-
ene, a thioxanthene antipsychotic, have also been shown to
inhibit MERS-CoV and SARS-CoV infection in vitro [38].
In addition, they were found to inhibit EVD VLP entry
[101]. Both are approved clinically, and clomipramine
belongs to the WHO Model List of Essential Medicines.
Several other neurotransmitter inhibitors, astemizole,
promethazine, chlorphenoxamine, and fluspirilene, were
active against MERS-CoV and SARS-CoV in cell culture,
but we were not able to find reports on activity against
other viruses [38]. Astemizole is an H1-histamine receptor
antagonist for treating allergic rhinitis that was withdrawn
from the market. Cardiac adverse events due to drug
overdose have been reported, but are extremely rare [105].
Recently, astemizole has gained renewed interest as an
anticancer and antimalarial drug [106, 107].
Chlorphenoxamine is an antihistamine and anticholin-
ergic that is currently in preclinical trials for malaria.
Fluspirilene is an approved antipsychotic for treatment of
schizophrenia. It is a known autophagy inducer [108].
Autophagy is a cellular degradative pathway that viruses
exploit for their propagation [109]. Modulators of autop-
hagy may perturb MERS-CoV or SARS-CoV infection,
and investigation of their broad-spectrum potential for the
treatment of coronaviral infections would be interesting
[110].
2.2.7 Nucleic Acid Synthesis Inhibitors
Several RNA/DNA synthesis inhibitors have broad-spec-
trum activity against viruses including SARS-CoV and
MERS-CoV [38, 80, 111–114]. Inosine monophosphate
dehydrogenase (IMPDH) inhibitors such as ribavirin,
mycophenolic acid, and mizoribine inhibit an important
step in de novo synthesis of nucleic acids although the
potency of these drugs against viruses varies. Ribavirin has
been used in combination with IFN in the clinic for treat-
ment of viral infections such as hepatitis C. Treatment
regimens with ribavirin are well characterized and have
been used in SARS and MERS patients with mixed results
(see 2.1). Ribavirin weakly inhibits MERS-CoV in vitro,
and conflicting data have been reported for the activity of
ribavirin against SARS-CoV [80, 111, 115]. Many of the
studies on ribavirin were performed in Vero cells that
reportedly have a defect in ribonucleoside uptake, which
could explain lack of activity for ribavirin in these cells
[116]. Another coronavirus, mouse hepatitis virus (MHV),
becomes sensitive to ribavirin when its exoribouclease
activity is inactivated. In presence of exoribonuclease,
ribavirin does not inhibit MHV replication [117]. The
MHV exoribonuclease has been suggested to function as a
‘proofreading’ viral enzyme that is necessary for high-fi-
delity replication of MHV. Similarly, the exoribonuclease
activity of MERS-CoV and SARS-CoV could possibly
counteract inhibitory activity of ribavirin.
Mycophenolic acid (MPA), an immunosuppressant used
to prevent organ rejection, has broad-spectrum antiviral
activities, and antifungal, antibacterial, anticancer, and
antipsoriatic properties [45, 118, 119]. Although MPA has
weak inhibitory activity against SARS-CoV in vitro, it has
promising activity against MERS-CoV [80, 112]. A
potential alternative to MPA, the prodrug mycophenolate
mofetil, has improved oral bioavailability [120].
Mycophenolate mofetil evaluated in the common mar-
moset model of MERS did not reduce disease manifesta-
tions compared to that observed in control subjects [121].
However, the MERS marmoset model does not recapitulate
human disease due to its rapid onset and pathology asso-
ciated with exposure methods [122]. Mizoribine, an
approved immunosuppressant in organ transplantation with
limited adverse side effects, has shown in vitro activity
against HCV and bovine viral diarrhea virus (BVDV), and
was considered as an alternative to ribavirin/IFN combi-
nations for treatment of HCV infections [123]. In vivo
analysis of ribavirin and other IMPDH inhibitors in SARS-
CoV-infected mice have suggested that these agents would
be of limited benefit [111].
The chemotherapeutic gemcitabine, has shown in vitro
activity against MERS-CoV and SARS-CoV [38]. The
drug’s anti-cancer mechanism is attributed to its ability to
inhibit ribonucleotide reductase essential for de novo
pyrimidine biosynthesis. Gemcitabine has been shown to
suppress influenza virus RNA transcription and replication
by targeting ribonucleotide reductase and showed anti-
retroviral activity in vivo in the mouse model for murine
leukemia virus [124, 125].
1950 J. Dyall et al.
2.2.8 Protease Inhibitors
MERS-CoV and SARS-CoV require activation of their
envelope glycoproteins by host proteases for cell entry by
the endosomal or the non-endosomal pathways. Inhibitors
of host cell proteases are being investigated as possible
antivirals [126]. The serine protease TMPRRSS2 mediates
entry via the non-endosomal pathway for both MERS-CoV
and SARS-CoV [50, 127–129]. Camostat mesylate, which
has been used in the treatment of chronic pancreatitis,
inhibits TMPSSR2-mediated glycoprotein activation of
MERS-CoV, SARS-CoV, and influenza virus
[126, 130–132]. K11777, a cysteine protease inhibitor, is in
clinical development for treating parasitic infections.
K11777 has broad-spectrum activity against coronaviruses
(MERS-CoV, SARS-CoV, HCoV-229E), filoviruses
(EBOV, Marburg virus), and paramyxoviruses (Nipah
virus) [133]. Interestingly, Zhou et al. [133] demonstrated
that Camostat and K11777 had inhibitory activity against
SARS-CoV whereas EBOV was only inhibited by K11777,
suggesting differential host protease requirements for these
viruses [133]. E-64-D, an inhibitor of an endosomal cys-
teine protease currently in Phase III trials for the treatment
of muscular dystrophy, inhibits both MERS-CoV and
SARS-CoV in vitro [38]. E-64-D also inhibits filovirus cell
entry [134, 135]. The dependency of viruses for specific
serine or cysteine host proteases must be considered in the
selection of protease inhibitors for antiviral therapeutic
applications. Therefore, an increased understanding of the
relationship between host proteases and viral pathogenesis
will determine the most effective treatment options for
viral infections.
Lopinavir was identified as an inhibitor of MERS-CoV
and SARS-CoV in vitro, and time-of-addition experiments
indicate that the drug acts at an early stage of viral entry
[36]. Lopinavir, an inhibitor of the HIV protease, is used
clinically for the treatment of HIV infections [136]. It is
given in combination with ritonavir, an inhibitor of cyto-
chrome P450 3A4, to increase blood concentrations
because of the low bioavailability of lopinavir [136].
Lopinavir also inhibits human papilloma virus and is cur-
rently under development for the topical treatment of cer-
vical cancer [137]. Treatment with lopinavir/ritonavir
resulted in reduced mortality in a NHP model of MERS
[121]. Lopinavir has been shown to target the main pro-
tease (Mpro) of SARS-CoV [138]. However, lopinavir has
also been shown to act on other intracellular processes that
are involved in coronavirus replication. Additional studies
are needed to fully understand the mechanism of action of
lopinavir involving cellular proteases. During the 2003
SARS outbreak, patients in open clinical trials were treated
with lopinavir/ritonavir in combination with ribavirin had a
milder disease course and reduction in fatality rate com-
pared to that observed with historical controls [27, 28].
2.2.9 Protein Synthesis Inhibitors
Three protein synthesis inhibitors with activity against
coronaviruses were identified, emetine, anisomycin and
omacetaxine mepesuccinate [38, 139]. Emetine, a natural
plant alkaloid, and anisomycin, an antibiotic, both inhibit
protein elongation and were identified as anti-protozoals
[140, 141]. While emetine is approved for amoebiasis
treatment, anisomycin did not move beyond clinical trials
[141, 142]. Dehydroemetine, a synthetic emetine deriva-
tive, has fewer side effects and is available as an investi-
gational new drug [143, 144]. Anisomycin was originally
discovered as a peptidyl transferase inhibitor, but also
activates the MAP kinase signaling pathway [145]. In
addition to activity against MERS-CoV and SARS-CoV,
emetine and anisomycin inhibit the animal picornavirus
encephalomyocarditis virus [146]. Anisomycin has in vitro
activity against poliovirus [147]. Omacetaxine mepesuc-
cinate, a plant-derived alkaloid, is an anticancer therapeutic
that received FDA approval in 2012 for the treatment of
chronic myeloid leukemia. Omacetaxine inhibits MERS-
CoV, bovine coronavirus, human enteric coronavirus and
hepatitis B virus [139, 148]. In spite of omacetaxine broad
spectrum anti-coronavirus activity, the drug had no activity
against SARS-CoV [38]. Drugs that inhibit coronaviruses
by targeting protein synthesis may have potential in the
development of combination therapies with drugs that
target other antiviral pathways.
2.2.10 Selective Estrogen Receptor Modulators
Recent investigations have demonstrated the potential of
estrogen receptor (ER) antagonists for repurposing as anti-
coronavirus compounds [38]. For example, toremifene
citrate and tamoxifen citrate with activity against SARS-
CoV and MERS-CoV were developed and approved as
anticancer therapeutics. Both drugs have shown activity
against HCV replication in vitro [149, 150]. Mechanistic
studies revealed that the ER is functionally associated with
HCV replication [150]. ER promotes the interaction
between the HCV replication complex and the HCV
polymerase NS5B. ER–mediated regulation of HCV gen-
ome replication is abrogated by tamoxifen.
Toremifene and tamoxifen also effectively inhibit
EBOV infection in vitro [151]. However, in contrast to
HCV, mechanistic studies have shown that toremifene-
mediated EBOV inhibition is independent of the ER
pathway as toremifene was still active against EBOV in
cells that did not express ER [151, 152]. Toremifene acts at
a late step of virus entry after internalization of EBOV and
Therapeutics for MERS-CoV and SARS-CoV 1951
may prevent fusion between the viral and endosomal
membranes [151–153]. Based on the chemical structure,
toremifene is a cationic amphiphilic drug (CAD) that is
known to be lysosomotropic and could affect endosomal
processes during virus entry [151, 154]. Treatment with
toremifene led to 50% survival of EBOV–infected mice
confirming that this drug has an effect in vivo as well
[151].
In terms of clinical application, toremifene and tamox-
ifen have good bioavailability, safety and tolerability pro-
files combined with a long history of use in the clinic.
However, prolongation of the QT interval has been noted
for toremifene and should not be prescribed to patients with
congenital or acquired long QT syndrome, uncorrected
hypokalemia or uncorrected hypomagnesemia [155].
Tamoxifen can increase uterine malignancies, stroke and
pulmonary embolism in women with ductal carcinoma
in situ or at high risk for breast cancer [156]. Despite these
side effects, the drugs may have substantial value for short-
term treatment of acute coronaviral infections. Advanced
patient studies and careful evaluation of the pharmacoki-
netic profiles may facilitate dosing strategies that limit the
risk of adverse events.
2.2.11 Sterol Metabolism Inhibitors
Two sterol synthesis inhibitors, terconazole and triparanol,
have shown activity against MERS-CoV and SARS-CoV
[38]. Studies with virus-like particles (VLPs) have
demonstrated that terconazole inhibits coronavirus cell
entry, including MERS-CoV and SARS-CoV. The sterol
synthesis pathway has been shown to be required for
infection by several viruses including HCV [157, 158].
Terconazole, approved for vaginal yeast infections, can be
administered orally, topically or by suppository. Triparanol
was approved for lowering plasma cholesterol, but was
withdrawn due to numerous side effects. Both are CADs
that induce accumulation of cholesterol in late endosomes
and have been shown to inhibit EBOV entry [152].
2.3 Drugs in Development
2.3.1 Potential Targets for Inhibition of MERS-CoV
and SARS-CoV
In addition to drug repurposing, development of novel
antiviral countermeasures is needed for emerging coron-
aviruses. To this end, design or development strategies
have targeted the viral replication cycle and host pathways
essential for viral replication (Table 4). Two nucleoside
inhibitors of viral RNA-dependent RNA polymerases, GS-
5734 and BCX4430, have potential as broad-spectrum
antivirals [159, 160]. Both drugs are active against MERS-
CoV and SARS-CoV in cell culture, but in vivo efficacy
remains to be investigated. In addition, a new class of
nucleosides with a flexible purine base has anti-coronaviral
activity, and further optimization could generate potent
inhibitors of the coronaviral polymerase [161]. The surface
glycoprotein (S) of SARS-CoV and MERS-CoV and other
coronaviruses consists of two domains: S1, containing the
receptor-binding domain (RBD) needed for extracellular
binding; and S2, containing the fusion peptide needed for
membrane fusion and release. Endocytosis of SARS-CoV
is facilitated by the binding of RBD with the angiotensin
converting enzyme 2 (ACE2) receptor on host cells.
Membrane-bound cathepsin L cleaves the S protein
revealing the S2 fusion protein, which fuses with the
membrane and releases the viral RNA. Inhibitors of
cathepsin L, the ACE2–SARS–S1 complex, or the S2
fusion peptide could be suitable targets to inhibit SARS-
CoV entry [162]. Results from recent studies have identi-
fied inhibitors of viral entry, viral proteases, and helicases
that potently inhibit both MERS-CoV and SARS-CoV
[162]. Proteases, such as papain-like protease and 3C-like
protease, could also be useful as antiviral targets for drug
development as they are required for cleaving non-struc-
tural proteins for viral maturation. Most protease inhibitors
are ‘‘suicide’’ protease inhibitors that bind to the target
irreversibly. However, reversible protease inhibitors may
have greater potential as they are less toxic and better
tolerated [113]. Recent studies with helicase inhibitors
show that three domain targets, N-terminal metal binding
domain, a hinge domain, and a NTP/helicase domain, have
potential for the development of new drugs [163].
2.3.2 RNA Interference
Directed RNA interference (RNAi) presents a powerful
approach for the development of novel virus-specific
therapeutics based on gene silencing [164]. Recent studies
have shown that small interfering RNAs (siRNAs) or short
hairpin RNAs can inhibit expression of viral genes and
thereby block the replication of SARS-CoV in cultured
cells [165–172]. Intranasal delivery of a combination of
small interfering RNA (siSC2-5) targeting SARS-CoV
open reading frame 1 and S protein decreased SARS
pathogenesis in NHPs [173]. Potential RNAi candidates
identified through computational modeling for MERS-CoV
require further in vitro and preclinical investigation [174].
Several RNAi therapeutics for the treatment of viral
infections have entered clinical trials including TKM-
Ebola, a siRNA/lipid nanoparticle platform targeting
EBOV [164, 175, 176]. This technology has great potential
for therapeutics for emerging viruses as viral genome
1952 J. Dyall et al.
sequencing can now be completed in a very short time
frame that is crucial in situations when an epidemic of a
novel emerging viral infection unfolds. The main obstacle
for RNAi strategies lies in the identification of suitable vi-
ral targets and in the delivery efficiency of nucleic acids to
target cells in vivo.
2.3.3 Peptide Entry Inhibitors
Peptides share common physicochemical properties with
CADs that facilitate interaction with cell membranes and
interference with the fusion of cellular and viral mem-
branes during virus entry. Researchers are making progress
in defining the mechanism of action of peptide entry
inhibitors of enveloped viruses such as enfuvirtide, an
approved HIV inhibitor [177]. Enfuvirtide is a 36-residue
peptide derived from the amphipathic loop/C-helix heptad
repeat domain of HIV gp41. A rational approach based on
scanning fusion protein sequences for amphipathic
sequences has led to the discovery of additional peptide
inhibitors for other viruses including MERS-CoV and
SARS-CoV [178–180]. Chemical modifications of peptides
have increased their in vivo stability and bioavailability,
improving their potential for clinical applications as novel
broad-spectrum viral entry inhibitors [181–183].
Table 4 Drugs in development for the treatment of Middle East respiratory syndrome (MERS) or severe acute respiratory syndrome (SARS)
Viral/cellular target Drug class Drug
MERS-CoV
3C-like protease Benzotriazole esters CE-5 [284]
Papain-like protease Thiopurines 6-Thioguanine, 6-mercaptopurine [113]
Helicase Triazole SSYA10-001 [162]
RNA-dependent RNA
polymerase
Nucleotide prodrug GS-5734 [159]
RNA-dependent RNA
polymerase
Nucleoside analog BCX4430 [160]
Membrane-bound
RNA synthesis
Small molecule inhibitor K22 [285]
Furin inhibitor Small molecule inhibitor Decanoyl-RVKR [286]
SARS-CoV
3C-like protease Benzotriazole esters CE-5 [284]
3C-like protease Anilides Peptide nitroanilides [287]
3C-like protease C2-symmetric inhibitors containing diol cores TL-3 [288]
3C-like protease Pyrazole analogs Pyrazolones [289]
3C-like protease Serine inhibitor Trifluoromethyl ketones [290]
3C-like protease Serotonin receptor antagonist Cinanserin [291]
3C-like protease Zinc-conjugated inhibitor JMF 1586 [292]
Papain-like protease Thiopurines 6-Thioguanine, 6-mercaptopurine [113]
Helicase Triazole SSYA10-001 [162]
Helicase Bananin derivatives Vanillinbananin, Idobananin [163]
NTPase/Helicase Aryl diketoacids Dihydroxychromone and
hydroxychromone derivatives
ADK analogs [293]; 2-(3-iodobenzyloxy)-6-(3-chlorobenzyloxy)-
5-hydroxychromone [294]
RNA-dependent RNA
polymerase
Nucleoside analogs BCX4430 [160], 4-aza-7,9-dideazaadenosine C-nucleosides [295],
fleximer nucleoside analogs [161]
Cathepsin L cellular
protease
Small molecule inhibitor Oxocarbazate [296], SSAA09E1 [297]
ACE2–SARS–S1
complex
Small molecule inhibitor SSAA09E2 [297]
S2-cell membrane
fusion
Small molecule inhibitor SSAA09E3 [297]
ACE2–SARS–S1
complex
Small molecule inhibitor NAAE [298]
ACE2 angiotensin converting enzyme, MERS-CoV Middle East respiratory syndrome coronavirus, S1 spike protein 1 domain, S2 Spike protein 2
domain
Therapeutics for MERS-CoV and SARS-CoV 1953
2.4 Antibody Therapy
The success of palivizumab for treatment of respiratory
syncytial virus infection has reinvigorated efforts to
develop monoclonal antibody-based therapeutics for
infectious diseases [184]. ZMapp, a monoclonal antibody
cocktail targeting EBOV, has been tested in NHPs with
success and was moved to Phase I and II clinical trials in
humans during the EVD epidemic [185, 186]. Similarly, a
monoclonal antibody against Hendra virus has been
administered to humans on a ‘‘compassionate use’’ basis
[187–189]. These examples demonstrate the potential for
antibody therapy to combat emerging/re-emerging viruses,
and similar strategies have been pursued by multiple
groups for development of antibodies to MERS-CoV
[190–197]. Monoclonal antibodies to MERS-CoV have
been sourced from humanized mice libraries or human
antibody libraries [192, 194, 197, 198]. Antibodies target
the S RBD, S1, or S2 subunits, and have demonstrated
efficacy in animal models as reviewed in Ying et al. [199].
Monoclonal antibody therapy can impart a selective pres-
sure for generation of resistant viruses. Although mutants
escaping monoclonal antibody pressure tend to be less fit,
analysis of the emergence of mutations that confer resis-
tance to the monoclonal antibody will need to be
performed.
An alternative to monoclonal antibody therapy is poly-
clonal antibody therapy using convalescent sera (sera
sourced from a nonhuman or humanized animal). Poly-
clonal antibodies provide an advantage over monoclonal
antibodies in that escape mutants are less likely to emerge
[200, 201]. Convalescent sera have been recommended for
MERS, and a Phase II interventional clinical trial is
ongoing to determine efficacy [198]. However, the avail-
ability of a suitable donor subject presents a significant
complication for this approach. Nonhuman animal sera has
been considered, but safety concerns limit this option.
Fractionation of nonhuman sera is an alternative; however,
antibody-mediated clearance can be limited due to failure
of the human Fc receptors to recognize the antibody heavy
chain.
An alternative is de-speciating the antibodies by using
only the Fab antibody fragment; however, cost and suffi-
cient material may make mass production of Fab fragments
difficult. The use of sera from humanized mice or other
small laboratory animals is complicated by sample acqui-
sition/volume restraints. Larger laboratory animals may
provide a potential alternative. SAB Biotherapeutics has
developed a trans-chromosomic bovine platform for the
generation of human IgG antibodies [201]. Vaccination of
trans-chromosomic cattle with S protein nanoparticles or
inactivated, whole virus generated fully humanized
polyclonal antibodies that demonstrated efficacy in the Ad-
DPP4 murine model of MERS.
3 Lessons Learned
One of the most important lessons regarding antiviral drug
development is that both highly specific and broad-spec-
trum antivirals bring unique advantages to the table.
Antiviral agents can fall anywhere in the spectrum between
‘‘broad-spectrum’’ and ‘‘highly specific’’ [202]. A drug that
targets a specific virus or virus family will have narrow
activity, high potency, and low toxicity; however, such a
drug may also promote resistance [202]. In contrast, broad-
spectrum antivirals typically target a host factor or path-
way, and often these agents have higher toxicities, lower
potencies, and delayed treatment effects. However, the
selective pressure for resistance is often lower with broad-
spectrum agents.
A large part of our knowledge on antiviral development
stems from the studies of chronic viral infections. Antiviral
development strategies for DNA viruses have been suc-
cessful in identifying a single drug that specifically targets
a viral protein. This strategy has been less successful for
RNA viruses. RNA viruses mutate at a higher rate than
DNA viruses resulting in enhanced development of drug
resistance.
3.1 AIDS
Despite extensive efforts over the past 30 years, a thera-
peutic or prophylactic HIV vaccine has remained elusive.
Antiviral agents are the only available treatments for AIDS.
Over 25 antivirals belonging to 6 different drug classes
targeting different stages of viral replication are available
(e.g. reverse transcriptase, protease, fusion, entry, inte-
grase) [203]. Combination treatments with 2 to 3 drugs are
effective and result in a sustained virologic response [204].
Two aspects have been found to be important for avoiding
resistance: (1) selecting drugs with at least two different
targets, and (2) selecting drugs that belong to different
chemical classes. These considerations may also apply for
drug combinations with synergistic effects against MERS-
and SARS-CoV.
3.2 Hepatitis C
Broad-spectrum antiviral therapies can be of great value for
treating emerging infections when it takes time to develop
direct-acting antivirals. For treatment of chronic hepatitis
C, clinicians have depended on IFN and ribavirin for a
number of years [205]. Eventually, IFN and ribavirin
combination was replaced by very effective fixed-
1954 J. Dyall et al.
combination therapies using direct-acting antivirals that
target multiple steps of the HCV life cycle [206]. IFN and
ribavirin contribute significantly to the treatment of viral
infections for which no direct-acting antivirals exist.
However, they have major side effects [207]. More options
for broad-spectrum antivirals with improved safety profiles
would be beneficial for use for emerging coronavirus
infections.
3.3 Influenza
Influenza viruses are characterized by a high mutation rate
of the RNA genome. As available vaccines may not be
protective against a novel pandemic strain, antiviral agents
are considered an essential component for preparedness.
Combinations of direct-acting antivirals are under evalua-
tion for additive or synergistic effects and prevention of
resistance [208, 209]. One triple combination (oseltamivir,
amantadine and ribavirin) is synergistic and prevents
resistance in vivo [210, 211], highlighting the potential of
combinatorial therapy.
3.4 Ebola Virus Disease
The recent epidemic of EVD inWestern Africa has renewed
the urgency for development of treatments against emerging
viruses. Although vaccines and direct-acting antiviral treat-
ment are under investigation, none are approved for clinical
use [212–216]. The WHO prioritized a panel of drugs
approved for other indications that were considered for
repurposing under FDA’s Emergency Use Authorization
[62]. Two of these drugs also have activity against MERS-
CoV and SARS-CoV, amodiaquine (antimalarial agent) and
toremifene citrate (breast cancer treatment).
Additional broad-spectrum antiviral agents (including
repurposed drugs) should be a top priority for future
emerging infections including coronavirus infections. A
panel of broad-spectrum drugs that have been carefully
validated for efficacy and safety and that could be used in
combination would supply a minimum of protection for
patients and healthcare workers at outbreak locations. This
panel of drugs could be used in situations of a known re-
emerging pathogen for which specific antiviral agents and
vaccines has not been approved or of an unknown novel
pathogen that could arise.
4 Gaps in Knowledge and Future Outlook
4.1 Animal Models of MERS and SARS
Effective development of countermeasures depends on
developing appropriate animal models that uniformly
recapitulate human disease progression and severity of
pathological manifestations. As with most animal models
of human disease, no one animal model fully reflects SARS
or MERS, therefore researchers are faced with exploring
several small animal models or choosing the ‘‘best-fit’’
model. To date, animal models do not fully recapitulate
human disease, thus animal models of MERS and SARS
need further refinement. Many small animal models have
been evaluated as potential MERS and SARS models
including mice, hamsters, and ferrets for MERS and Syrian
hamsters, and guinea pigs for SARS [217–219]. Four
murine models have been reported for MERS. The first
model that demonstrated promise involved transduction of
the respiratory tract with the putative MERS-CoV receptor,
human DPP4 (or CD26) [220]. The major indicator of
disease in this model is viral load in the lung at 4 days post-
inoculation. Although clinical signs of disease, including
weight loss, were limited in this model, it has been used for
pathogenesis countermeasure studies [221, 222]. Lethal,
disseminated MERS infection has been demonstrated in
transgenic mice expressing human DPP4 [223]. Inflam-
matory processes were observed in the brains of these mice
in contrast to human disease in which CNS involvement
has not been reported. A transgenic mouse MERS model
was developed in which the mouse DPP4 gene was
replaced by the human DPP4 gene under control of the
endogenous mouse DPP4 promoter. Using this model,
MERS-CoV-infected mice developed lung pathology
[194]. In addition, administration of human monoclonal
antibodies against the spike protein in these transgenic
mice provided protection against MERS-CoV infection
[194]. Clustered regularly interspaced short palindromic
repeat-CRISPR-associated protein 9 (CRISPR-CAS9) gene
editing technology was used to modify the mouse DPP4 to
match human DPP4 by altering amino acids at positions
288 and 330. Interestingly, wild type virus infection of
these mice did not result in an improved model of MERS.
However, serial passage of MERS-CoV resulted in MERS-
CoV-15. Intranasal exposure of MERS-CoV-15 in
288/330, mice led to weight loss and a severe respiratory
disease that included ARDS-like signs and reduced pul-
monary function [224].
MERS-CoV infection of rabbits has also been evaluated
as a model for MERS. Haagmans et al. [225] demonstrated
that MERS-CoV infected rabbits did not develop obvious
clinical signs, but infectious virus could be detected in the
upper respiratory tract [225]. Furthermore, epithelial cells
of the bronchioles and terminal bronchioles respiratory
tract were positive for MERS-CoV by immunohistochem-
istry and in-situ hybridization, which reflects tissue tropism
in human disease [225]. Using the rabbit model, Houser
et al. [191] demonstrated that human monoclonal antibody
336 given pre-exposure reduced viral RNA lung titer at
Therapeutics for MERS-CoV and SARS-CoV 1955
3 days post-exposure, but not when given post-exposure
[191].
Due to phylogenetic similarities with humans, NHP
models of disease have long been considered as necessary
for evaluation of countermeasures to infectious diseases.
Rhesus monkey and common marmoset have been evalu-
ated as potential models for MERS. Following intratracheal
instillation of MERS-CoV in rhesus monkey models, lung
pathology was observed [122, 226–228]. Experiments
using rhesus monkeys have indicated that they develop
limited systemic disease and a transient respiratory disease.
Radiologic evaluations have indicated inflammatory infil-
trates that develop shortly after exposure. Analysis of lung
tissues by reverse transcriptase- quantitative polymerase
chain reaction indicated virus replication in the lung.
Similar to MERS, African green monkeys (AGMs),
rhesus monkeys, cynomolgus monkeys, and common
marmosets have been identified as potential models for
SARS [229]. Smits et al. [230] compared SARS CoV
infection in young AGMs to cynomolgus monkeys, they
observed that neither species developed clinical signs
during a 4-day experiment [230]. Gross pathology indi-
cated multi-focal pulmonary consolidation with consoli-
dated grey-red firm lungs. These lesions affected 30% of
the lungs in one subject. By comparison, the cynomolgus
monkeys developed small patchy macroscopic lesions.
Similar to MERS-CoV infection of NHPs, viral load
decreases from exposure day to day 4. A comparison
between AGMs, rhesus monkeys, and cynomolgus mon-
keys further support AGMs as the best available NHP
model for SARS [231]. AGMs developed the highest viral
load and most disease when compared to cynomolgus and
rhesus monkeys. Lethal disease was not observed in any of
these species; therefore, further development of the SARS
model is warranted since lethal respiratory tract disease
was the hallmark of SARS.
As an alternative to Old World NHPs, many groups have
employed marmosets as models of human infectious dis-
ease [232–234]. Common marmosets have been evaluated
as a MERS model [121, 122, 235]. These studies have
demonstrated that common marmosets develop disease
following exposure to MERS-CoV as shown by
histopathological analysis, radiological analysis, and RT-
qPCR. However, variable results have been reported, and
exposure methodology can impact disease progression.
Therefore, mock-infected groups must be included to
account for pathological artifacts. The virus-specific
pathology could be quantified using computed tomography,
and future experiments using large group sizes could be
used for countermeasure evaluation. Greenough et al. [236]
performed a serial euthanasia study of SARS-CoV infected
marmosets [236]. Subjects were intratracheally exposed
with SARS-CoV. They observed mild inconsistent clinical
signs of disease. Viral loads peaked at day 4 post-infection.
Histopathology indicated interstitial pneumonitis with
multinucleated syncytia that were described as mild and
not observed in all late time-point subjects. Overall, further
research is needed to develop animal models of SARS that
reflect human disease presentation.
4.2 Combinations with Synergy
Drugs with repurposing potential discussed here (Table 3)
have the advantage of easy access, availability and
decreased cost of development and provide a wide array of
options for combination studies. The pharmacological
knowledge available for such compounds may also reduce
concerns regarding adverse effects in patients. The gener-
ation of a translational database encompassing pharmaco-
dynamics data and infectious disease biology data has been
proposed and would greatly facilitate decision making to
pursue new drug combinations [237]. Many of the drugs
have potential for broad-spectrum antiviral activity and
have already been in clinical use for treating other viral
infections. As novel drugs in development move from the
pre-clinical to clinical phase, they also become available
for combination therapy. Care should be taken with the
pharmacological evaluation of each combination to avoid
possible contraindications of the drugs with regards to
disease or adverse effects. Novel broad-spectrum replica-
tion inhibitors, such as GS-5734 (Gilead Sciences, in Phase
I clinical trial), immunomodulators (nitazoxanide; steroids;
statins) along with direct-acting antiviral agents for coro-
naviruses that are in development represent interesting
partners for combinations. Combinations can involve
broad-spectrum versus specific antiviral agents; drugs with
different mechanism of action; or drugs that target different
steps of the viral life cycle. Identifying one or more potent
combinations with activity in an animal model would
greatly increase preparedness for the next coronavirus
outbreak.
4.3 Structure-Based Drug Design
Elucidation of the crystal structure of viral proteins has led
to novel approaches for rational drug design. Rational
design investigations using protein structure information
and in silico screening for affinity to active sites of viral
proteins holds promise. HIV-1 protease inhibitors have
been one of the big successes of rational drug design. Only
6 years after the publication of the HIV-1 protease struc-
ture, saquinavir was developed in record time from bench
to bedside and was licensed for use against AIDS in 1995
[238–240]. In total, six antivirals against the HIV-1 pro-
tease were designed and approved between 1995 and 2000.
Similarly, for HCV, computer–aided approaches based on
1956 J. Dyall et al.
the known crystal structure of a viral protein have suc-
cessfully guided the design and synthesis of inhibitors for
the HCV NS3/NS4A proteases such as the peptidomimet-
ics, telaprevir and boceprivir [241–243].
Due to the power of computational modeling using
crystal structures from known coronaviruses, the crystal
structures for the viral proteases, Mpro and PLpro, of SARS-
CoV and MERS-CoV were determined relatively quickly
[244]. These structures have already been used for the
discovery of inhibitors with high binding affinity to the
active site of the proteases. The structures of additional
MERS-CoV or SARS-CoV viral proteins have yet to be
determined and would offer additional viral targets for drug
discovery.
Structural design can help with development of inhi-
bitors in preparing for future outbreaks of yet unknown
emerging coronaviruses. Based on the potential of zoo-
notic transmission of coronaviruses from bats to humans,
crystal structures for the main protease (Mpro) of dif-
ferent bat coronavirus families have been proposed for
screening and identifying broad-spectrum antiviral
agents [244]. Proof-of-principle was shown for the novel
protease inhibitor SG85, which inhibits the bat coron-
avirus HKU4 [244]. In 2012, SG85 was quickly identi-
fied as an inhibitor of MERS-CoV through in silico
docking studies with the Mpro of MERS-CoV and the bat
coronavirus HKU4.
4.4 Cationic Amphiphilic Drugs: A Novel Class
of Antiviral Agents?
Several drugs (e.g. toremifene citrate, terconazole,
Fig. 3) belong to a group of compounds termed cationic
amphiphilic drugs (CADs) [100, 151, 152, 154, 245].
Phenothiazines (e.g. fluphenazine chlorpromazine,
Fig. 3) are CADs that have been shown to inhibit HCV
entry at virus-host cell fusion by intercalating into the
cholesterol-rich domains of the host cell membrane and
increasing membrane fluidity [97]. Drugs that act
through this mechanism may present an interesting new
class of broad-spectrum antivirals. CADs are known to
be lysomotropic and accumulate in acidic compartments
where their tertiary amine groups are protonated. The
compounds act as mild bases and can neutralize the low
pH of the acidic environment of endo/lysosomes. CADs
can intercalate into membranes, alter the biophysical
properties of membranes and thereby could potentially
interfere with fusion of virus with the endo/lysosomal
membrane. The concept of interfering with virus entry
and budding through physicochemical properties of
drugs is intriguing. Many viruses would be susceptible to
this type of inhibition and CADs could be used as broad-
spectrum antiviral agents. Detailed structure-activity
relationship studies on CADs will be required to deter-
mine the chemical core structures and physicochemical
properties important for this type of antiviral agent.
Future investigations regarding the conservation of this
mechanism of action to coronaviruses, as well as other
emerging viruses, are warranted.
4.5 Analogs of Developed Drugs
Approved drugs with activity against MERS-CoV or
SARS-CoV could be used as lead compounds for further
antiviral drug development. Pharmaceuticals usually have
undergone multiple rounds of structure-activity relation-
ship studies generating analogs to improve drug activity
against the original indication or target. The analogs of
drugs with activity against MERS-CoV and SARS-CoV
may still be available and could be screened to identify
other analogs with increased antiviral activity. Although
drug analogs would have to go through the full licensure
process, there would be little to no added initial cost
associated with producing the structure-activity relation-
ship compounds. Recycling old analogs is one approach
that may have value for developing novel drugs for viral
infections.
5 Conclusion
A more streamlined process is needed for development of
effective treatment measures for emerging and re-emerging
pathogens. The availability of a panel of approved broad-
spectrum drugs would clearly be beneficial as they could be
used for treating disease symptoms and reducing morbidity
until more specific acting antivirals and vaccines are
developed.
A large number of potential drugs and therapeutics for
the treatment of MERS and SARS have been discussed.
The greatest challenge will be how best to down-select and
evaluate the different approaches. As we have learned from
drug development for AIDS and hepatitis, alleviating dis-
ease symptoms and increasing life span may be a more
achievable goal rather than looking for a treatment that will
provide complete recovery. Broad-spectrum antivirals,
specific antivirals, and immune modulators each have an
important role in treating viral infections either individu-
ally or in combination. Effective communication between
the different institute partners (government, industry, aca-
demic; national, and international partners) is essential.
Combining these drug discovery efforts will increase the
chance of having one or more potential therapeutic agents
at an advanced development stage by the time another
outbreak of an emerging coronavirus occurs.
Therapeutics for MERS-CoV and SARS-CoV 1957
Acknowledgements The authors thank Laura Bollinger and Jiro
Wada for providing technical writing services and graphical support,
respectively.
Compliance with Ethical Standards
The content of this publication does not necessarily reflect the views
or policies of the US Department of Health and Human Services
(DHHS) or of the institutions and companies affiliated with the
authors.
Funding This work was supported in part by the Division of Intra-
mural Research and Division of Clinical Research, National Institute
of Allergy and Infectious Diseases. This work was funded in part
through Battelle Memorial Institute’s prime contract with the US
National Institute of Allergy and Infectious Diseases (NIAID) under
Contract No. HHSN272200700016I. J.D. performed this work as
employee of Tunnell Government Services, Inc., subcontractor to
Battelle Memorial Institute (BMI). R.G. performed this work as
employee of BMI. G.G.O. performed this work as employee of MRI
Global, subcontractor to BMI.
Conflict of interest The authors have declared that they have no
conflict of interest.
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